Author

Defense Date

2006

Document Type

Thesis

Degree Name

Master of Science

Department

Biochemistry

First Advisor

Dr. Gregorio Gil

Abstract

Bile acids regulate their own synthesis through a feedback regulatory mechanism of mainly two enzymes in the classic pathway, the 7α-hydroxylase and the 12α-hydroxylase. In the early 1990's it was shown that the regulatory responses of 7α-hydroxylase are mediated at the transcriptional level and since then many positive and negative transcription factors that mediate regulatory response have been identified. An important finding was that the transcription factors regulating the expression of 7α- and 12α-hydroxylase genes are nuclear receptors.One of the first nuclear receptors identified to play a role in the transcription of the 7α-hydroxylase gene was HNF-4 since then many nuclear receptors have been identified that are involved in regulating the 7α- and 12α-hydroxylase genes. Among them the most important ones are FTF and HNF-4 which has been shown to play crucial roles in the transcription and regulation by bile acids. In this study we demonstrate the importance of FTF and HNF-4 independent of each other in the transcription and bile acid-mediated regulation of the 7α- and 12α-hydroxylase enzymes by creating promoter mutants that would either bind FTF or HNF- 4. Once the binding studies were established we performed tissue culture experiments to confirm the promoter activity and bile acid-mediated regulation with the respective promoter mutant constructs. The data from this study shows that HNF-4 is important for 7α-hydroxylase promoter activity but is not required and importantly we show that HNF-4 is not a required for the bile acid-mediated regulation of the 7α-hydroxylase. We present data which suggests that FTF is absolutely required for the promoter activity and bile acid-mediated regulation of 7α-hydroxylase. With respect to the 12α-hydroxylases how that both FTF and HNF-4 are absolutely required for promoter activity. In this study we present evidence that since the bile acid responsive elements (BARE) are similar within both the 7α- and 12α-hydroxylase promoters one can be exchanged for the other maintaining both activity and bile acid-mediated regulation.